The goal of this research is to develop advanced magnetic resonance spectroscopy (MRS) and imaging techniques and to apply them and other complementary methods to studying brain metabolism, neurotransmission, and enzyme activity. MRS allows in vivo measurement of the neurotransmission of glutamate and GABA, which plays important roles in many major psychiatric diseases, including depression and schizophrenia. During 2018-2019 we further developed our 7 Tesla proton MRS technique for converting glutamate, glutamine and GABA intro pseudo singlets at 56 ms echo time by introducing a radiofrequency pulse during the evolution of the strongly coupled spins. Taking advantage of the greatly enhanced glutamate H4 and glutamine H4 signals, we have successfully performed , for the first time, in vivo experiments for detecting carbon-13 labeling of glutamate and glutamine in human brain at 7 Tesla using standard hardware and at the sensitivity and spatial resolution of proton MRS. Furthermore, we have found a way to greatly improve experimental determination of macromolecule baseline by variable inversion recovery. Our method is able to capture the main spectral features of macromolecule resonances seen in the metabolite-nulled spectrum. This technique has significantly improved spectral quantification for proton MRS at medium echo time as it uses both longitudinal and transverse relaxation differences to resolve macromolecule baseline from metabolite signals (An et al, Determination of Macromolecule Baselines by Variable Inversion-Recovery of Metabolites, Proceedings of the International Society for Magnetic Resonance in Medicine, P. 2243 (2019)). To investigate the detrimental effect of carrier frequency mismatch on spectral editing experiments we used full density matrix simulation and Monte Carlo analysis to assess how editing yields are affected by various extents of carrier frequency mismatch. Surprisingly, we found significant errors in metabolite quantification without frequency matching of basis functions when carrier frequency mismatch was generally considered negligible. By matching basis functions with the history of frequency deviation, errors in glutamate, glutamine, amma-aminobutyric acid, and glutathione concentrations were reduced to negligible levels (An et al, Effects of Carrier Frequency Mismatch on Frequency-Selective Spectral Editing, Proceedings of the International Society for Magnetic Resonance in Medicine, P. 512 (2019); An et al, Effects of carrier frequency mismatch on frequency-selective spectral editing, Magn. Reson. Mater. Phy., 32:237-246 (2019)). Therefore, we recommend that all spectral editing experiments should correct for carrier frequency mismatch caused by patient movement and system instability. Building on our recent development of a novel technique for measuring transverse relaxation of metabolites using radiofrequency pulse driven steady state (Li et al, A novel approach to probing in vivo metabolite relaxation: Linear quantification of spatially modulated magnetization, Magn. Reson. Med., 79:2491-2499 (2018); L. Li et al, Systems and methods for probing in vivo metabolite relaxation by linear quantification of spatially modulated magnetization, International Patent Application No. PCT/US2018/054340; U.S. Provisional Patent Application No. 62/567,991 (2018)), we have developed an improved echo time-independent technique for measuring transverse relaxation time and demonstrated that this radiofrequency-driven longitudinal steady state technique can reliably measure glutamate transverse relaxation in the frontal cortex, where structural and functional abnormalities have been associated with psychiatric symptoms. Because glutamate resides predominantly in glutamatergic neurons and its relaxation properties reflect the intracellular environment of glutamatergic neurons we will push for applying our method to characterize the intracellular environment of glutamatergic neurons in a variety of brain disorders. The improved method is capable of reliably measuring glutamate relaxation from human subjects in a typical clinical setting (Li et al, Quantification of in vivo transverse relaxation of glutamate in the frontal cortex of human brain by radio frequency pulse-driven longitudinal steady state. PLoS One. 2019 Apr 17;14(4):e0215210. doi: 10.1371/journal.pone.0215210). In addition to developing the above cell-type-specific MRS techniques we have also made significant progress in expanding the scope of MRS by increasing the sensitivity of MRS signals.